CHRONIC HYPOXIA AND RESISTANCE TO MYOCARDIAL ISCHEMIA

说明(申请人提供)：血小板生成素(TPO)是通过TPO受体MPL调节巨核细胞和血小板发育的主要生理调节因子。最近，我们偶然发现TPO是一种非常有效的心脏保护剂。这些原始的观察结果可能会导致我们对TPO的机制和生物学效应的看法发生范式转变。因此，这项建议的总体目标是确定TPO提供早期和延迟心脏保护的细胞内信号通路。虽然有大量的文献关于MPL受体的组成、分布和在血小板功能中的信号转导，但还没有关于TPO与MPL结合提供心脏保护的途径以及这一途径如何被心脏中的低氧招募的信息。我们假设TPO受体的激活触发了促生存通路，并抑制了细胞凋亡，从而通过开放线粒体KATP通道和/或线粒体来提供早期和延迟的心脏保护！KCA频道。具体地说，我们将：1)确定TPO提供早期心脏保护的途径。我们将确定TPO信号是否通过其受体(MPL)激活促生存激酶信号级联：Jak/STAT、RAS、Raf、磷脂酰肌醇-3-羟基激酶(PI3K)/Akt、PKC、JNK、p38Mark和p42/44Mark。我们将确定TPO在再灌注后立即给予的保护是否通过激活有利于生存的风险途径起作用，以及TPO是否保护慢性缺氧的心脏。2)确定线粒体KATP通道、Kca通道和肌膜KATP通道在TPO早期心肌保护中的相对重要性。我们将确定早期心肌保护中的TPO信号是否由线粒体KATP和KCA通道的开放所介导。我们将测量TPO处理的心脏线粒体的线粒体膜电位、呼吸和ATP合成，以确定KATP通道与线粒体功能的关系。3)研究慢性低氧对兔和人心脏TPO及其受体(MPL)表达的影响。我们将利用我们的慢性缺氧模型，在婴儿、人和兔的心脏上确定是否对慢性缺氧的适应增加了MPL的信息和蛋白的表达以及血小板的循环水平。4)确定TPO产生的延迟性心肌保护介质。我们将确定“NO是否作为TPO延迟性心脏保护的媒介。然后我们将确定TPO是否通过开放肌膜和/或线粒体KATP通道来调节延迟性心脏保护。这些研究结果有望影响我们对TPO在心脏中生物学的理解，并可能导致治疗心肌梗死的新疗法的发展。

DESCRIPTION (provided by applicant): Thrombopoietin (Tpo) is the primary physiologic regulator of megakaryocyte and platelet development through the Tpo receptor mpl. Recently we have made the novel and serendipitous observation that Tpo is an extraordinarily potent cardioprotective agent. These original observations may cause a paradigm shift in how we view the mechanisms and biological effects of Tpo. Thus the overall objective of this proposal is to define the intracellular signaling pathways by which Tpo confers early and delayed cardioprotection. Whereas there is a substantial body of literature regarding the mpl receptor composition, distribution and signaling in platelet function, there is no information about the pathway by which binding of Tpo to mpl confers cardioprotection and how this pathway is recruited by hypoxia in the heart. We hypothesize that activation of the Tpo receptor triggers pro-survival pathways and suppresses apoptosis to confer early and delayed cardioprotection by opening of mitochondrial KATp channels and/or mitochondria! KCa channels. Specifically we shall: 1) Determine the pathways that confer early cardioprotection by Tpo. We shall determine whether Tpo signals through its receptor (mpl) to activate the pro-survival kinase signaling cascade: JAK/STAT, Ras, Raf, phosphatidylinositol-3-OH kinase (PI3K)/Akt, PKC, JNK, p38 MARK and p42/44 MARK. We shall determine whether protection by Tpo given immediately following reperfusion acts by activating the pro-survival RISK pathway and whether Tpo protects the chronically hypoxic heart. 2) Determine the relative importance of the mitochondrial KATP channel, KCa channel and the sarcolemmal KATP channel in early cardioprotection produced by Tpo. We shall determine if Tpo signaling in early cardioprotection is mediated by opening of mitochondrial KATP and KCa channels. We shall measure mitochondrial membrane potential, respiration and ATP synthesis in mitochondria isolated from Tpo-treated hearts to determine the relationship between KATP channels to mitochondrial function. 3) Determine the effect of chronic hypoxia on the expression of Tpo and the Tpo receptor (mpl) in rabbit and human heart. We shall utilize our model of chronic hypoxia to determine in both infant human and rabbit hearts whether adaptation to chronic hypoxia increases expression of message and protein for mpl and circulating levels of platelets. 4) Determine the mediators of delayed cardioprotection produced by Tpo. We shall determine if "NO acts as a mediator of delayed cardioprotection by Tpo. We shall then determine if mediation of delayed cardioprotection by Tpo occurs by opening sarcolemmal and/or mitochondrial KATP channels. The results of these studies are expected to have an impact on our understanding of the biology of Tpo in heart and may lead to the development of new therapeutics for the treatment of myocardial infarction.